Increased generation of reactive oxygen species (ROS) and altered redox homeostasis are important biochemical characteristics of cancer cells, and play a major role in cancer cell survival and drug sensitivity. Our recent study showed that chronic lymphocytic leukemia (CLL) cells exhibited intrinsic oxidative stress and were highly dependent on cellular antioxidants such as glutathione (GSH) to maintain redox balance. We also found that that were highly dependent on the bone marrow stromal cells to alleviate the oxidative stress and promote their survival and drug resistance through enhancing GSH synthesis mediated by an intercellular metabolic communication involving cystine->cysteine conversion and shuttling between the two cell compartments. These research findings provide new mechanistic insights into the biochemical interaction between leukemia and stromal cells. Importantly, the dependency of CLL cells on stromal cells for redox balance and cell survival has revealed an Achilles heel of CLL cells that can be targeted for therapeutic purpose. This grant renewal application will build on these novel findings, and will test the hypothesis that stimulation of GSH synthesis in CLL cells by the bone marrow stromal cells through the metabolic communication between the two cell types is essential for CLL cell survival and drug resistance, and that abrogation of this protective mechanism may represent a novel therapeutic strategy to effectively kill CLL cells in the stromal environment and overcome drug resistance in vivo. The specific aims of this project are (1) to investigate the biochemical basis for the high dependency of CLL cells on bone arrow stromal cells to maintain GSH synthesis and promote cell survival and drug resistance, (2) to develop new therapeutic strategies to target the critical survival mechanism of CLL cells by abolishing the stromal-induced GSH protection of leukemia cells aiming at effectively killing CLL cells in the presence of bone marrow stromal cells, and (3) to test the in vivo therapeutic activity of new strategies to abolish the stromal protection of CLL cells using novel agents that target GSH metabolism in combination with standard anti-CLL drugs in a novel CLL mouse model. This study is expected to significantly advance our understating of leukemia-stromal interaction by elucidating a key redox-modulating mechanism that promotes leukemia cell survival and drug resistance. This study will also lead to the development of novel therapeutic strategies targeting the Achilles heel of CLL by abolishing the stromal protection. Since the stromal microenvironment plays a major role in promoting CLL cell survival and drug resistance and currently there are very few strategies that can effectively overcome stromal protection of CLL cells, the results of our study will likely have significant therapeutic implications in clinical treatment of this most common adult leukemia. PUBLIC HEALTH RELEVANCE: Increased generation of reactive oxygen species (ROS) and redox alterations are important biochemical properties of cancer cells and play a major role in cancer cell survival and drug resistance. The main objectives of this research are to test the hypothesize that promotion of GSH synthesis in chronic lymphocytic leukemia (CLL) cells by the bone arrow stromal cells through the metabolic communication between the two cell compartments is essential for CLL cell survival and drug resistance, and that abrogation of this stromal protection will render CLL cells highly vulnerable to drug treatment in vivo. This study will reveal a novel mechanism of stromal-leukemia interaction that is therapeutically important, and will develop new strategies to target the Achilles heel of CLL to improve in vivo therapeutic activity.